Human T-cell leukemia virus type I (HTLV-I) is the etiologic agent of adult T-cell leukemia (ATL). ATL is a complex and multi-faceted disease that evolves initially from viral-induced carcinogenic events and leads to multiple chromosomal abnormalities that are associated with the aggressive subtypes of ATL. Tax, the viral oncoprotein encoded within the pX region of the viral genome, is considered a major contributor to cell cycle deregulation in HTLV-I transformed cells by targeting such cellular factors as cyclin D2, p21/waf1, p16/INK4a, and p53. This concerted deregulation, especially at the G1 phase of the cell cycle, ensures continuous cellular progression via the loss of checkpoint control and insensitivity to anti-mitogens. The long-term goal of our research is to understand how Tax-dependent perturbation of restriction point regulators leads to leukemogenesis. A significant regulator of the G1 restriction point in eukaryotic cells is the tumor suppressor protein retinoblastoma (Rb). Rb, with p107 and p130, are part of a family of proteins involved in quiescence, cellular division, differentiation, senescence, and apoptosis. We have recently found that within both HTLV-I transformed cells and ATL patients there are a decrease of Rb protein as compared to uninfected controls. Specifically, this decrease is dependent on Tax expression and regulated at the posttranslational level. We have found that Tax is able to bind to Rb within the B domain through either a LXCXE or PENF homology motif within the C terminus of Tax. We further show, through in vitro degradation assays, that Rb degradation is dependent on Tax's ability to associate with the 26S proteasome. The proteasome degradation of Rb appears to be restricted to the hypophosphorylated (or active) Rb species, since the predominant Rb species remaining in HTLV-I transformed cells is the inactive form of Rb. Therefore, loss of the active species of Rb has led us to speculate that this results in a phenotype that is similar to cells that have lost Rb activity due to genetic/epigenetic alterations. Our hypothesis is that Rb degradation by Tax contributes to the immortalization of HTLV-I infected cells. Our rationale for these studies is based on data that shows the HTLV-I viral protein Tax destabilizes the Rb protein by targeting this protein to the 26S proteasome for degradation. We believe this mechanism is analogous to HPV's E7-dependent destabilization of Rb that results in the loss of the active form of Rb. Rb deregulation within the p16/INK4a/cyclin D pathway is an early event in HPV-16 immortalization by promoting cell cycle entry and proliferation of senescent primary cells. These phenotypes are also observed in HTLV-I transformed cells as shown by our preliminary data and may contribute to ATL malignancy. The following specific aims will address our hypothesis: (I) what is the mechanism of Tax-dependent proteolysis of Rb in HTLV-I transformed cells? (II) Is degradation of Rb an early event that contributes to Tax-dependent immortalization? Data obtained from these studies will shed light on how Tax and Rb interaction leads to the immortalization of HTLV-I infected ATL cells.